The basic objective of this research program is to develop specific information about the spectrum of mutations that can be induced by environmental carcinogens. We propose to study, at the nucleotide level, the DNA alterations that are generated by carcinogens in the lacI gene of Escherichia coli. We will determine the mutagenic spectrum induced by benzo(a)pyrene, 2-acetylaminofluorene, aflatoxin B1, and ultra-violet light in bacteria with different DNA repair capabilities. The information we expect to obtain from these studies includes: 1) the full range of mutational events that arise from damage to DNA by selected environmental carcinogens: 2) the influence of certain genes governing the enzymatic repair of DNA lesions on the mutagenic spectrum of carcinogens: 3) the effect of primary structure (sequence) on the conversion of DNA lesions to mutations. To develop this information, we shall induce and map mutations in the lacI gene carried on an F' factor. The mutation will then be crossed onto a plasmid whose copy number can be amplified. The isolated plasmid will be cut with a restriction enzyme to remove the region containing the lacI gene. The 5' ends of this fragment will be labelled with 32P and the labelled DNA will be cut with a set of restriction enzymes to generate fragments that are suitable for sequencing by the Maxam-Gilbert method. To permit analysis of the mutagenic potential of ultraviolet light induced DNA lesions we shall damage a segment of the lacI gene with UV light in vitro to introduce blocks to replication. These segments will be analyzed for the distribution of damage on the one hand and inserted into a phage lambda vector on the other hand. The modifications which ensue subsequent to DNA repair will be determined by sequencing the lacI insert in plaque forming phage.